This project will test the general hypothesis that mitotic misregulation is the key determinant in chondrocyte aging. In the first phase, we propose to examine the transcript profiles of chondrocytes, and identify those genes whose expression changes with age, and osteoarthritis utilizing comprehensive DNA microarrays that cover more than 90% of the encoded genes in the human genome. A database will be established to manage this large amount of information and provide access to the public in a web-based interactive format. Some of the proposed capabilities will include hierarchical clustering, promoter analysis, and automated batch search to all public databases, including our comprehensive human and mouse transcriptome database. The second phase of this project will focus on functional analyses. Both computer-assisted sequence search and biochemical assays will be utilized to define the functional roles of genes whose expression changes with age and severity of osteoarthritic pathology. We expect a large proportion of genes identified in this study to be expressed-sequence tags (ESTs). We propose to develop high-throughput systems for cloning and protein expression, along with assays for functional characterization. Specific Aim 1: Examine the transcript profiles of chondrocytes in full thickness normal human articular cartilage from donors age 20-90, with at least 10 samples per decade. Specific Aim 2: Obtain transcript profiles of microdissected OA cartilage to distinguish early from advanced disease and fibrillated, calcified and fibrocartilage. Specific Aim 3: Establish a comprehensive database for articular cartilage aging and osteoarthritis development and provide access to the public in a web- based interactive format. Specific Aim 4: Clone differentially expressed ESTs. Specific Aim 5: Characterize gene functions in assays for chondrocyte proliferation and survival, differentiation, extracellular matrix remodeling, and inflammation. The proposed studies have the potential to identify genetic changes that determine joint aging and thus define new molecular markers for diagnosis and targets for prevention and therapeutic intervention.